Demyelinating diseases are disorders concerning the myelin sheaths of the nervous system. Myelin sheaths, which cover many nerve fibers, are composed of lipoprotein layers formed in early life. Myelin is formed by the oligodendroglia in the CNS and promote transmission of a neural impulse along an axon.
Many congenital metabolic disorders (e.g. phenylketonuria and other aminoacidurias; Tay-Sachs, Niemann-Pick, and Gaucher's diseases; Hurler's syndrome; Krabbe's disease and other leukodystrophies) affect the developing myelin sheath, mainly in the CNS. Unless the biochemical defect can be corrected or compensated for, permanent, often widespread, neurological deficits result.
Demyelination in later life is a feature of many neurological disorders; it can result from damage to nerves or myelin due to local injury, ischemia, toxic agents, or metabolic disorders. Extensive myelin loss is usually followed by axonal degeneration and often by cell body degeneration, both of which may be irreversible. However, remyelination occurs in many instances, and repair, regeneration, and complete recovery of neural function can be rapid. Recovery often occurs after the segmental demyelination that characterizes many peripheral neuropathies; this process may account for the exacerbations and remissions of multiple sclerosis (MS). Central demyelination (i.e. of the spinal cord, brain, or optic nerves) is the predominant finding in the primary demyelinating diseases, whose etiology is unknown. The most well known demyelinating disease is MS (see below).
Further demyelinating diseases comprise:
Acute disseminated encephalomyelitis, which is characterized by perivascular CNS demyelination, and which can occur spontaneously but usually follows a viral infection or viral vaccination;
Acute inflammatory peripheral neuropathies that follow a viral vaccination or the Guillain-Barré syndrome, they affect only peripheral structures;
Adrenoleukodystrophy and adrenomyeloneuropathy, which are rare X-linked recessive metabolic disorders characterized by adrenal gland dysfunction and widespread demyelination of the nervous system;
Leber's hereditary optic atrophy and related mitochondrial disorders, which are characterized primarily by bilateral loss of central vision, and which can resemble the optic neuritis in MS; and
HTLV-associated myelopathy, a slowly progressive spinal cord disease associated with infection by the human T-cell lymphotrophic virus, that is characterized by spastic weakness of both legs.
Multiple sclerosis (MS) is a slowly progressive CNS disease characterized by disseminated patches of demyelination in the brain and spinal cord, resulting in multiple and varied neurological symptoms and signs, usually with remissions and exacerbation (see The Merck Manual Home Edition, www.merck.com).
The cause is unknown but an immunological abnormality is suspected, with few clues presently indicating a specific mechanism. Postulated causes include infection by a slow or latent virus, and myelinolysis by enzymes. IgG is usually elevated in the CSF, and elevated titers have been associated with a variety of viruses, including measles. The significance of these findings and of reported associations with HLA allotypes and altered number of T cells is unclear, and the evidence somewhat conflicting. An increased family incidence suggests genetic susceptibility; women are somewhat more often affected than men. Environmental factors seem to be present. Although age at onset generally is from 20 to 40 years, MS has been linked to the geographic area where a patient's first 15 years are spent. Relocation after age 15 does not alter the risk.
Plaques or islands of demyelination with destruction of oligodendroglia and perivascular inflammation are disseminated through the CNS, primarily in the white matter, with a predilection for the lateral ad posterior columns (especially in the cervical and dorsal regions), the optic nerves, and periventricular areas. Tracts in the midbrain, pons, and cerebellum also are affected, and gray matter in both cerebrum and cord may be affected.
Cell bodies and axons are usually preserved, especially in early lesions. Later, axons may be destroyed, especially in the long tracts, and a fibrous gliosis gives the tracts their “sclerotic” appearance. Both early and late lesions may be found simultaneously. Chemical changes in lipid and protein constituents of myelin have been demonstrated in and around the plaques.
The disease is characterized by various symptoms and signs of CNS dysfunction, with remissions and recurring exacerbations. The most common presenting symptoms are paresthesias in one or more extremities, in the trunk, or on one side of the face; weakness or clumsiness of a leg or hand; or visual disturbances, e.g., partial blindness and pain in one eye (retrobulbar optic neuritis), dimness of vision, or scotomas. Other common early symptoms are ocular palsy resulting in double vision (diplopia), transient weakness of one or more extremities, slight stiffness or unusual fatigability of a limb, minor gait disturbances, difficulty with bladder control, vertigo, and mild emotional disturbances; all indicate scattered CNS involvement and often occur months or years before the disease is recognized.
The course is highly varied, unpredictable, and, in most patients, remittent. Life span is probably not shortened except in the most severe cases. At first, months or years of remission may separate episodes, especially when the disease begins with retrobulbar optic neuritis. Remissions can last >10 years. However, some patients have frequent attacks and are rapidly incapacitated; for a few, particularly for male patients with onset in middle age, the course can be rapidly progressive. Exposure to excess heat from fever or the environment sometimes worsens symptoms.
Diagnosis is indirect, by deduction from clinical and laboratory features. MRI, the most sensitive diagnostic imaging technique, may show plaques. Gadolinium-contrast enhancement can distinguish areas of active inflammation from older brain plaques. MS lesions may also be visible on contrast-enhanced CT scans, in which sensitivity may be increased by giving twice the iodine dose and delaying scanning (double-dose delayed CT scan).
CSF is abnormal in the majority of patients. IgG may be >13%, and lymphocytes and protein content may be slightly increased. Oligoclonal bands, which indicate IgG synthesis within the blood-brain barrier, may be detected by agarose electrophoresis of CSF in up to 90% of patients with MS, but absence of these bands does not rule out MS. IgG levels correlate with disease severity. Myelin basic protein may be elevated during active demyelination.
Spontaneous remissions and fluctuating symptoms make treatments difficult to evaluate. Corticosteroids are the main form of therapy. They may shorten the symptomatic period during attacks, although they may not affect eventual long-term disability. Patients presenting with acute severe optic neuritis may delay the onset of MS by using high-dose IV corticosteroids.
Immunosuppressive drugs (methotrexate, azathioprine, cyclophosphamide, cladribine) are generally used for more severe progressive forms. Immunomodulatory therapy with interferon-β reduces the frequency of relapses in MS. Other promising treatments still under investigation include other interferons, oral myelin, and glatiramer to help keep the body from attacking its own myelin. Glatiramer is a synthetic co-polymer with similarities to myelin basic protein and is administered by daily subcutaneous injection. Its main action is thought to be suppression of the immune response against myelin to promote immune tolerance (Clegg and Bryant, 2001).
Interferons are cytokines, i.e. soluble proteins that transmit messages between cells and play an essential role in the immune system by helping to destroy micro-organisms that cause infection and repairing any resulting damage. Interferons are naturally secreted by infected cells and were first identified in 1957. Their name is derived from the fact that they “interfere” with viral replication and production.
Interferons exhibit both antiviral and antiproliferative activity. On the basis of biochemical and immunological properties, the naturally-occurring human interferons are grouped into three major classes: interferon-alpha (leukocyte), interferon-beta (fibroblast) and interferon-gamma (immune). Alpha-interferon is currently approved in the United States and other countries for the treatment of hairy cell leukemia, venereal warts, Kaposi's Sarcoma (a cancer commonly afflicting patients suffering from Acquired Immune Deficiency Syndrome (AIDS)), and chronic non-A, non-B hepatitis.
Further, interferons (IFNs) are glycoproteins produced by the body in response to a viral infection. They inhibit the multiplication of viruses in protected cells. Consisting of a lower molecular weight protein, IFNs are remarkably non specific in their action, i.e. IFN induced by one virus is effective against a broad range of other viruses. They are however species-specific, i.e. IFN produced by one species will only stimulate antiviral activity in cells of the same or a closely related species. IFNs were the first group of cytokines to be exploited for their potential anti-tumor and antiviral activities.
The three major IFNs are referred to as IFN-α, IFN-β and IFN-γ. Such main kinds of IFNs were initially classified according to their cells of origin (leukocyte, fibroblast or T cell). However, it became clear that several types may be produced by one cell. Hence leukocyte IFN is now called IFN-α, fibroblast IFN is IFN-β and T cell IFN is IFN-γ. There is also a fourth type of IFN, lymphoblastoid IFN, produced in the “Namalwa” cell line (derived from Burkitt's lymphoma), which seems to produce a mixture of both leukocyte and fibroblast IFN.
The interferon unit or International unit for interferon (U or IU, for international unit) has been reported as a measure of IFN activity defined as the amount necessary to protect 50% of the cells against viral damage. The assay that may be used to measure bioactivity is the cytopathic effect inhibition assay as described (Rubinstein, et al. 1981; Familletti, P. C., et al., 1981). In this antiviral assays for interferon about 1 unit/ml of interferon is the quantity necessary to produce a cytopathic effect of 50%. The units are determined with respect to the international reference standard for Hu-IFN-beta provided by the National Institutes of Health (Pestka, S. 1986).
Every class of IFN contains several distinct types. IFN-β and IFN-γ are each the product of a single gene.
The proteins classified as IFNs-α are the most diverse group, containing about 15 types. There is a cluster of IFN-α genes on chromosome 9, containing at least 23 members, of which 15 are active and transcribed. Mature IFNs-α are not glycosylated.
IFNs-α and IFN-β are all the same length (165 or 166 amino acids) with similar biological activities. IFNs-γ are 146 amino acids in length, and resemble the α and β classes less closely. Only IFNs-γ can activate macrophages or induce the maturation of killer T cells. In effect, these new types of therapeutic agents can be called biologic response modifiers (BRMs), because they have an effect on the response of the organism to the tumor, affecting recognition via immunomodulation.
In particular, human fibroblast interferon (IFN-β) has antiviral activity and can also stimulate natural killer cells against neoplastic cells. It is a polypeptide of about 20,000 Da induced by viruses and double-stranded RNAs. From the nucleotide sequence of the gene for fibroblast interferon, cloned by recombinant DNA technology, (Derynk et al. 1980) deduced the complete amino acid sequence of the protein. It is 166 amino acid long.
Shepard et al. (1981) described a mutation at base 842 (Cys→Tyr at position 141) that abolished its anti-viral activity, and a variant clone with a deletion of nucleotides 1119-1121.
Mark et al. (1984) inserted an artificial mutation by replacing base 469 (T) with (A) causing an amino acid switch from Cys→Ser at position 17. The resulting IFN-β was reported to be as active as the ‘native’ IFN-β and stable during long-term storage (−70° C.).
REBIF (recombinant human interferon-β) is a recent development in interferon therapy for multiple sclerosis (MS) and represents a significant advance in treatment. REBIF is interferon(IFN)-beta 1a, produced from mammalian cell lines. It was established that interferon beta-1a given subcutaneously three times per week is efficacious in the treatment of Relapsing-Remitting Multiple Sclerosis (RR-MS). Interferon beta-1a can have a positive effect on the long-term course of MS by reducing number and severity of relapses and reducing the burden of the disease and disease activity as measured by MRI (The Lancet, 1998).
Tumor Necrosis Factor, or TNF, previously called Cachectin, is a pleiotropic cytokine released by activated T cells and macrophages. TNF is a member of the interferon, interleukin and colony stimulating factor cytokine network, which has a key role in signaling with regard to the pathogenesis of many infectious and inflammatory diseases by inducing a number of proinflammatory changes, including production of other cytokine and adhesion molecule (Fiers, 1991).
For convenience, the term TNF collectively shall mean, in the entire context of the present application, both Tumor Necrosis Factor-alpha or Tumor Necrosis Factor-beta from animals or humans, together with naturally occurring alleles thereof (Pennica et al., 1984, Wallach et al., 1986, Beutler, B. and Cerami, A. (1987)). TNF-beta, also called lymphotoxin, has a similar activity but is produced by different cell types (lymphocytes and Natural Killer cells) in response to antigenic or mitogenic stimuli (Gray et al., 1984).
Thus, Tumor Necrosis Factor (TNF-α) and Lymphotoxin (TNF-β) are cytokines which have many effects on cells. Some of their effects are likely to be beneficial to the organism: they may destroy, for example, tumor cells or virus infected cells and augment antibacterial activities of granulocytes. In this way, TNF contributes to the defense of the organism against infectious agents and to recovery from injury. But both TNF-α and TNF-β have also been described to have deleterious effects. There is evidence that overproduction of TNF-α can play a major pathogenic role in several diseases. In some diseases, TNF may cause excessive loss of weight (cachexia) by suppressing activities of adipocytes and by causing anorexia and TNF-α was thus called cachectin. It was also described as a mediator of the damage to tissues in rheumatic diseases and as a major mediator of the damage observed in graft-versus-host reactions.
TNF is expressed as a mature 17 kDa protein that is active as a trimer. This complex exerts its biological activity by aggregating their cell surface receptors, which mediate specific effects in different organs and tissues.
TNF exerts its activity, which is required for the normal development and function of immune system, by binding a family of membrane bound receptor molecules including p55 TNF receptor I, defined in the literature also TNF-RI, and p75 TNF receptor, defined in the literature also TNF-RII (Bazzoni and Beutler, 1996). The dominance of TNF-RI in transducing TNF signal is suggested by the ability of agonistic antibodies specific for this receptor to mimic the majority of TNF induced responses (Shalaby et al., 1990). By binding to its membrane-bound receptors, TNF triggers the signaling pathway through cytoplasmic mediators like TRADD and TRAP-1 (for TNF-RI) or TRAF-1 and TRAF-2 (for TNF-RII), leading to different cell response, like T cell proliferation, tumor-cell lysis in vitro, dermal necrosis, insuline resistance, apoptosis. The extracellular portions of both TNF receptors can be shed and these soluble receptors retain the ability to bind TNF, inactivating TNF activity by formation of high affinity complexes, thereby reducing the binding of TNF to target cell membrane receptors (Nophar et al., 1990).
Based on the finding that TNF-alpha immunoreactivity has been found in high levels in MS lesions, TNF has been described to play a role in the pathogenesis of multiple sclerosis (Darlington, 1999). Therefore, it was generally thought that TNF should be blocked or reduced in order to treat MS, and TNF blocking agents have been suggested for treatment of multiple sclerosis (Selmaj et al., 1995). However, experiments using mice lacking TNF, so-called TNF −/− mice, showed that these mice developed severe neurological impairment with extensive inflammation and demyelination upon induction of a MS like disease with the protein MOG (Liu et al., 1998).
Truncated forms of the TNF-RI (p55) and TNF-RII (p75) receptors mentioned above are described e.g. in EP914431. These soluble receptors are called TBPI and TBPII, respectively (Engelmann et al., 1990). The natural and recombinant soluble TNF receptor molecules, and methods of their production have been described e.g. in the European Patents EP 308 378, EP 398 327 and EP 433 900. EP 398 327 describes that TBPs are not only inhibitors of TNF activity, but also maintain the beneficial effect of TNF. It has also been described that the soluble TNF-receptors stabilize the bioactivity of TNF and thus augment some of its effects (Aderka et al., 1992).
A TNF-like activity was also shown for antibodies directed against the soluble forms of the TNF-receptors (Engelmann et al., 1990).
In addition to this, EP 880 970 discloses the use of TBPs for treatment of multiple sclerosis.